# US 9,517,264 B2 · Human FGF receptor and β-Klotho binding proteins

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- Number: US 9,517,264 B2
- Document ID: US9517264B2
- Status: Granted
- Kind: B2
- Application number: 13/641,041
- Filed: 2011-04-13
- Published: 2013-05-23
- Granted: 2016-12-13
- Earliest claimed: 2010-04-15
- Assignees: Amgen Inc.
- Applicants: Roger Fachini; Ian Foltz; Seog Joon Han; Susie Miki Harris; Shaw-Fen Sylvia Hu; Chadwick Terence King; Yang Li; Ji Lu; Mark Leo Michaels; Jeonghoon Sun
- Inventors: Roger Fachini; Ian Foltz; Seog Joon Han; Susie Miki Harris; Shaw-Fen Sylvia Hu; Chadwick Terence King; Yang Li; Ji Lu; Mark Leo Michaels; Jeonghoon Sun
- Web page: https://patentgrep.com/patent/US9517264B2

## CPC

- A61K 39/3955: {against proteinaceous materials, e.g. enzymes, hormones, lymphokines}
- A61P 1/16: for liver or gallbladder disorders, e.g. hepatoprotective agents, cholagogues, litholytics
- A61P 3/00: Drugs for disorders of the metabolism (of the blood or the extracellular fluid A61P7/00)
- A61P 3/04: Anorexiants; Antiobesity agents
- A61P 3/06: Antihyperlipidemics
- A61P 3/10: for hyperglycaemia, e.g. antidiabetics
- A61P 9/00: Drugs for disorders of the cardiovascular system
- A61P 43/00: Drugs for specific purposes, not provided for in groups A61P1/00-A61P41/00
- C07K 16/2863: {against receptors for growth factors, growth regulators}
- C07K 16/40: against enzymes
- C07K 2317/21: from primates, e.g. man
- C07K 2317/31: multispecific
- C07K 2317/32: specific for a neo-epitope on a complex, e.g. antibody-antigen or ligand-receptor
- C07K 2317/75: Agonist effect on antigen
- C07K 2317/92: Affinity (KD), association rate (Ka), dissociation rate (Kd) or EC50 value
- C07K 2319/75: containing a fusion for activation of a cell surface receptor, e.g. thrombopoeitin, NPY and other peptide hormones
- A61K: PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES (devices or methods specially adapted for bringing pharmaceutical products into particular physical or administering forms A61J3/00; chemical aspects of, or use of materials for deodorisation of air, for disinfection or sterilisation, or for bandages, dressings, absorbent pads or surgical articles A61L; soap compositions C11D)
- A61P: SPECIFIC THERAPEUTIC ACTIVITY OF CHEMICAL COMPOUNDS OR MEDICINAL PREPARATIONS
- C07K: PEPTIDES (peptides in foodstuffs A23; obtaining protein compositions for foodstuffs, working-up proteins for foodstuffs A23J; preparations for medicinal purposes A61K; peptides containing beta-lactam rings C07D; cyclic dipeptides not having in their molecule any other peptide link than those which form their ring, e.g. piperazine-2,5-diones, C07D; ergot alkaloids of the cyclic peptide type C07D519/02; macromolecular compounds having statistically distributed amino acid units in their molecules, i.e. when the preparation does not provide for a specific; but for a random sequence of the amino acid units, homopolyamides and block copolyamides derived from amino acids C08G69/00; macromolecular products derived from proteins C08H1/00; preparation of glue or gelatine C09H; single cell proteins, enzymes C12N; genetic engineering processes for obtaining peptides C12N15/00; compositions for measuring or testing processes involving enzymes C12Q; investigation or analysis of biological material G01N33/00)

## Abstract

The present invention provides compositions and methods relating to or derived from antigen binding proteins and antigen binding protein-FGF21 fusions that specifically bind to β-Klotho, or β-Klotho and one or more of FGFR1c, FGFR2c, FGFR3c, and FGFR4. In some embodiments the antigen binding proteins and antigen binding protein-FGF21 fusions induce FGF21-like signaling. In some embodiments, an antigen binding protein or antigen binding protein-FGF21 fusion antigen binding component is a fully human, humanized, or chimeric antibody, binding fragments and derivatives of such antibodies, and polypeptides that specifically bind to β-Klotho, or β-Klotho and one or more of FGFR1c, FGFR2c, FGFR3c, and FGFR4. Other embodiments provide nucleic acids encoding such antigen binding proteins and antigen binding protein-FGF21 fusions, and fragments and derivatives thereof, and polypeptides, cells comprising such polynucleotides, methods of making such antigen binding proteins and antigen binding protein-FGF21 fusions, and fragments and derivatives thereof, and polypeptides, and methods of using such antigen binding proteins and antigen binding protein-FGF21 fusions, fragments and derivatives thereof, and polypeptides, including methods of treating or diagnosing subjects suffering from type 2 diabetes, obesity, NASH, metabolic syndrome and related disorders or conditions.

## Claims

### Claim 1

An antigen binding protein comprising:
(a) a light chain variable domain comprising; (i) a light chain CDR1 comprising an amino acid sequence of SEQ ID NO: 107; (ii) a light chain CDR2 comprising an amino acid sequence of SEQ ID NO: 113; (iii) a light chain CDR3 comprising an amino acid sequence of SEQ ID NO: 121; and
(b) a heavy chain variable domain comprising: (i) a heavy chain CDR1 comprising an amino acid sequence of SEQ ID NO: 84; (ii) a heavy chain CDR2 comprising an amino acid sequence of SEQ ID NO: 90; and (iii) a heavy chain CDR3 comprising an amino acid sequence of SEQ ID NO: 99; wherein the antigen binding protein specifically binds β-Klotho.

### Claim 2 (depends on 1)

The isolated antigen binding protein of claim 1, wherein the antigen binding protein comprises: (a) a light chain variable domain sequence comprising SEQ ID NO: 18; and (b) a heavy chain variable domain sequence comprising SEQ ID NO: 29; wherein the antigen binding protein specifically binds to β-Klotho.

### Claim 3 (depends on 2)

The antigen binding protein of claim 2, wherein the antigen binding protein comprises: (a) the light chain constant sequence of SEQ ID NO: 13; (b) the light chain constant sequence of SEQ ID NO:15; (c) the heavy chain constant sequence of SEQ ID NO: 9; or (d) the light chain constant sequence of SEQ ID NO: 13 or SEQ ID NO: 15 and the heavy chain constant sequence of SEQ ID NO: 9.

### Claim 4 (depends on 1)

The antigen binding protein of claim 1, wherein the antigen binding protein comprises a human antibody, a humanized antibody, chimeric antibody, a monoclonal antibody, a polyclonal antibody, a recombinant antibody, an antigen-binding antibody fragment, a single chain antibody, a diabody, a triabody, a tetrabody, a Fab fragment, an F(fa′)x fragment, a domain antibody, an IgD antibody, an IgE antibody, an IgM antibody, an IgG1 antibody, an IgG2 antibody, an IgG3 antibody, an IgG4 antibody, or an IgG4 antibody having at least one mutation in the hinge region.

### Claim 5 (depends on 1)

The antigen binding protein of claim 1, that, when bound to β-Klotho: (a) binds to β-Klotho with substantially the same Kd as a reference antibody; (b) induces FGF21-like signaling of 10% or greater than the signaling induced by a wild-type FGF21 standard comprising the mature form of SEQ ID NO:2 as measured in an ELK-luciferase reporter assay; (c) exhibits an EC50 of 10 nM or less of FGF21-like signaling in an assay selected from the group consisting of: (i) a FGFR1c/β-Klotho-mediated in vitro recombinant cell-based assay; (d) exhibits an EC50 of less than 10 nM of agonistic activity on FGFR1c in the presence of β-Klotho in an in vitro recombinant FGFR1c receptor mediated reporter assay; and (e) an EC50 of greater than 1 μM of agonistic activity on FGFR1c in the absence of β-Klotho in an in vitro recombinant FGFR1c receptor mediated reporter assay; and (f) competes for binding with a reference antibody to β-Klotho, wherein the reference antibody comprises one or more of the following combinations of light chain and heavy chain variable domain sequences: L1H1, L2H2, L3H3, L4H4, L5H5, L6H6, L7H7, L8H8, L9H9, L10H10 and L11H11.

### Claim 6 (depends on 5)

The antigen binding protein of claim 5, that, when bound to β-Klotho: (a) lowers blood glucose in an animal model; (b) lowers serum lipid levels in an animal model; or (c) (a) and (b).

### Claim 7 (depends on 1)

A pharmaceutical composition comprising the antigen binding protein of claim 1 in admixture with a pharmaceutically acceptable carrier thereof.

### Claim 8 (depends on 1)

An isolated nucleic acid comprising a polynucleotide sequence encoding the light chain variable domain, the heavy chain variable domain, or both, of the antigen binding protein of claim 1.

### Claim 9

An isolated nucleic acid comprising a polynucleotide sequence encoding, SEQ ID NO: 18, SEQ ID NO: 29, or both.

### Claim 10 (depends on 9)

An expression vector comprising the nucleic acid of claim 9.

### Claim 11 (depends on 9)

An isolated cell comprising the nucleic acid of claim 9.

### Claim 12 (depends on 11)

The isolated cell of claim 11, comprising an expression vector comprising the nucleic acid.

### Claim 13 (depends on 12)

A method of producing an antigen binding protein that specifically binds to β-Klotho comprising incubating the host cell of claim 12 under conditions that allow it to express the antigen binding protein.

### Claim 14 (depends on 1, 2)

An antigen binding protein of any of claims 1, 2, and 3-6, wherein the heavy chain comprises a peptide that specifically binds to one or more of FGFR1c, FGFR2c, FGFR3c, and FGFR4.

### Claim 15 (depends on 14)

A heavy chain of the antigen binding protein of claim 14, wherein the peptide comprises one or more of: TRLWKYWV (SEQ ID NO: 184); RRLYIFWE (SEQ ID NO: 185); YKAWGYYV (SEQ ID NO: 186); YQAWGYYV (SEQ ID NO: 187); YQAWGYLV (SEQ ID NO: 188); YQAWGYFV (SEQ ID NO: 189); FTWVFWNV (SEQ ID NO: 190); YQVWGYFV (SEQ ID NO: 191); YKWLKWNL (SEQ ID NO: 192); RRLYIFEW (SEQ ID NO: 193); WAERGG (SEQ ID NO: 194); GGWAVGRI (SEQ ID NO: 195); YKYLVFWV (SEQ ID NO: 196); YKYLSYWV (SEQ ID NO: 197); YKTAWYWK (SEQ ID NO: 198); YVFHKWWV (SEQ ID NO: 199); YVFYLWWK (SEQ ID NO: 200); YRWLHWHV (SEQ ID NO: 201); YKFLFWHA (SEQ ID NO: 202); RRQWGFWV (SEQ ID NO: 203); YSAWSFWV (SEQ ID NO: 204); LARWGFWV (SEQ ID NO: 205); YDAWGYWV (SEQ ID NO: 206); WRKYYHFWVS (SEQ ID NO: 207); KRLYGLFWYD (SEQ ID NO: 208); KKHWSSLFFE (SEQ ID NO: 209); KAWPYSWEAV (SEQ ID NO: 210); EWYCGVLFNCQQ (SEQ ID NO: 211); HFGCGVIFNCVSD (SEQ ID NO: 212); WELCASGYGWCYLH (SEQ ID NO: 213); APSCKSYIGFGLYHCWDG (SEQ ID NO: 214); and or HFKCGMGLFECADP (SEQ ID NO: 215).

### Claim 16 (depends on 15)

The antigen binding protein heavy chain of claim 15, wherein the heavy chain comprises a CH2 loop, a CH3 loop or both a CH2 and a CH3 loop.

### Claim 17 (depends on 16)

The heavy chain of claim 16, wherein the heavy chain comprises a CH3 loop.

### Claim 18 (depends on 17)

The heavy chain of claim 17, wherein the CH3 loop comprises the peptide.

### Claim 19 (depends on 16)

The heavy chain of claim 16, wherein the heavy chain comprises a CH2 loop.

### Claim 20 (depends on 19)

The heavy chain of claim 19, wherein the CH2 loop comprises the peptide.

### Claim 21 (depends on 15, 18, 20)

An antigen binding protein comprising the heavy chain of claim 15, 18 or 20.

### Claim 22 (depends on 14)

A pharmaceutical composition comprising the antigen binding protein of claim 14 in admixture with a pharmaceutically acceptable carrier thereof.

### Claim 23 (depends on 14)

An isolated nucleic acid comprising a polynucleotide sequence encoding the light chain variable domain, the heavy chain variable domain, or both, of the antigen binding protein of claim 14.

### Claim 24 (depends on 23)

The isolated nucleic acid of claim 23, comprising a polynucleotide sequence encoding SEQ ID NO: 18, SEQ ID NO: 29, or both.

### Claim 25 (depends on 24)

An expression vector comprising the nucleic acid of claim 24.

### Claim 26 (depends on 24)

An isolated cell comprising the nucleic acid of claim 24.

### Claim 27 (depends on 26)

The isolated cell of claim 26, comprising an expression vector comprising the nucleic acid.

### Claim 28 (depends on 27)

A method of producing an antigen binding protein that specifically binds to β-Klotho comprising incubating the host cell of claim 27 under conditions that allow it to express the antigen binding protein.

### Claim 29 (depends on 1)

An antigen binding protein-FGF21 fusion comprising: (a) an antigen binding component, wherein the antigen binding component comprises an antigen binding protein of claim 1; and (b) an FGF21 component.

### Claim 30 (depends on 29)

The antigen binding protein-FGF21 fusion of claim 29, wherein the FGF21 component comprises at least 25 consecutive residues of SEQ ID NO:341.

### Claim 31 (depends on 30)

The antigen binding protein-FGF21 fusion of claim 30, wherein the FGF21 component comprises (a) SEQ ID NO:342 or (b) SEQ ID NO:343.

### Claim 32 (depends on 29)

The antigen binding protein-FGF21 fusion of claim 29, further comprising a linker.

### Claim 33 (depends on 29, 31)

The antigen binding protein-FGF21 fusion of claim 29 or claim 31, wherein the antigen binding component comprises SEQ ID NOS:18 and 29.

### Claim 34 (depends on 32)

The antigen binding protein-FGF21 fusion of claim 32, wherein the linker is selected from the group consisting of (G4S)3, (SEQ ID NO: 336) (G4S)6 (SEQ ID NO: 337), (G4S)9 (SEQ ID NO: 338), (G4S)12 (SEQ ID NO: 339) and (G4S)15 (SEQ ID NO: 340).

### Claim 35 (depends on 29)

The antigen binding protein-FGF21 fusion of claim 29, wherein the FGF21 component is joined to the heavy chain of the antigen binding component.

### Claim 36 (depends on 35)

The antigen binding protein-FGF21 fusion of claim 35, wherein the heavy chain comprises one or more of: SEQ ID NOs:316, 320, 322, 324, 326, 318, 328, 330, 332 and 334.

### Claim 37 (depends on 29)

The antigen binding protein-FGF21 fusion of claim 29, wherein the FGF21 component is joined to the light chain of the antigen binding component.

### Claim 38 (depends on 29)

The antigen binding protein of claim 29, that, when bound to β-Klotho, β-Klotho and one or more of FGFR1c, FGFR2c, FGFR3c, and FGFR4: (a) lowers blood glucose in an animal model; (b) lowers serum lipid levels in an animal model; or (c) (a) and (b).

### Claim 39 (depends on 29)

A pharmaceutical composition comprising an antigen binding protein-FGF21 fusion of claim 29, further comprising a pharmaceutically acceptable carrier.

### Claim 40 (depends on 7, 22)

A method of treating a condition in a subject in need of such treatment comprising administering a therapeutically effective amount of the composition of claim 7, 22, or 39 to the subject, wherein the condition is treatable by lowering blood glucose.

### Claim 41 (depends on 40)

The method of claim 40, wherein the condition is selected from type 2 diabetes, obesity, dyslipidemia, NASH, cardiovascular disease, and metabolic syndrome.